This group has focused on the conserved homeobox DNA binding domain as a means to gain insight into the molecular basis of early embryonic development in mammals. Homeobox genes occupy key positions in the regulatory gene hierarchy responsible for establishing the embryonic body plan in Drosophila, and are thought to play analogous roles in the development of higher vertebrates. Several novel homeobox genes have been previously isolated by the laboratory that are expressed in early embryogenesis. Current research has concentrated on the Rpx gene, which possesses the most anterior-restricted expression domain of all known homeobox genes. Transcripts are seen very early in the cephalic neural plate, and become progressively restricted to Rathke's pouch, the rudiment of the anterior pituitary, indicating a potential role for the Rpx gene in the development of this organ. Experiments to determine how this gene is regulated in the embryo and how it functions in development are in progress. Chimeric fusion genes consisting of varying amounts of the 5' flanking sequence of the gene linked to the bacterial beta-galactosidase (lacZ) reporter gene have been used to generate transgenic mice. Analysis of fusion gene expression in transgenic embryos indicates that all of the information sufficient for proper temporal and spatial expression resides within 1 kb of upstream sequence. The sequence of this region is characterized by the presence of several motifs found in the regulatory regions of other genes expressed in the anterior pituitary, at least one of which is conserved between the mouse and frog homologs of the Rpx gene. More precise deletion or mutation of these regions is underway in order to more accurately dissect the elements necessary for proper expression. Determination of elements specifically required for Rpx expression will potentially lead to the identification of genes that occupy earlier positions in a regulatory hierarchy at work in the early embryo. Experiments designed to investigate the function of Rpx include generating gain-of-function and loss-of-function models by ectopically expressing Rpx gene products in transgenic mice and by mutating the gene through homologous recombination in embryonic stem cells.
